The invention relates generally to equipment used to analyze chemical and biological samples in multiwell containers, commonly called microplates. Currently, they have up to 1536 wells in an array measuring about 3.4 by 5 inches (87xc3x97127 mm). Many types of analysis are possible, each one employing a light detector to determine the amount of light emitted from a sample in one of the wells. In general, analyses or measurements are made by fluorescence, absorbance, or luminescence. Most analyses involve measuring the light emitted in response to excitation light directed into the sample or as the result of the introduction of chemical reagents. The present invention provides a means for carrying out each of these general types of analysis in a single instrument, a commercial embodiment of which is designated the Fusion(trademark) Universal Microplate Analyzer by its manufacturer, the Packard Instrument Company.
Other analyzers which are able to carry out similar measurements on chemical and biological samples are disclosed in a large number of patents. Representative of more recent patents are the analyzers discussed in the following.
U.S. Pat. No. 6,042,785 assigned to Wallac OY shows an instrument capable of performing various spectrographic measurements. Two types of excitation light sources are used and two detectors for receiving light emitted from the sample, either passing through a mirror or through an aperture.
LJL Bio Systems in U.S. Pat. Nos. 6,097,028 and 6,071,748 shows a multifunctional analyzer which employs a plurality of excitation light sources and emitted light detectors. Optical switches are included to direct the excitation light and emission light to and from the sample. A feature of the instrument is its ability to limit the light to a xe2x80x9csensed volumexe2x80x9d away from the walls of the sample container.
Lab Systems OY in U.S. Pat. Nos. 6,144,455 and 6,084,680 shows a fluorometer in which a partly reflective mirror has a plurality of areas transparent to excitation and emitted light and a plurality of areas which are non-transparent to excitation and emitted light.
The present inventors have sought to develop a multi-purpose analyzer capable of carrying out various types of measurements in an effective and efficient manner. Their universal microplate analyzer is described below.
In one aspect, the invention is a universal microplate analyzer, as shown in the accompanying illustrations and described below. The analyzer includes the following features:
Analyses which can be carried out on samples include fluorescence, absorption, and luminescence.
Measurements may be made in at least four modes:
excitation light is introduced into the sample well from the top and light is emitted from the top and directed to a light detector;
excitation light is introduced into the sample well from the bottom and light emitted from the bottom is directed to a light detector;
light is introduced through the bottom of the well and light which is not absorbed leaves through the top and is directed to a light detector;
no light is introduced to the sample well and light generated by chemical reaction leaves through the top of the well and is directed to a light detector.
Multiple excitation light sources can be included. Two types of excitation light sources are used in a preferred embodimentxe2x80x94a flash light source e.g. a flash Xenon arc lamp, and a continuous wave light source e.g. a quartz tungsten halogen lamp. The light from each source is filtered to provide the desired spectral band for the intended analysis and thereafter the filtered light is shaped and directed to a sample well.
Excitation light is directed via optical fiber channels to enter a sample containing well through either the top via a read head or to the bottom, depending on the optical fiber channel which is selected.
One of several types of optical devices is switched into position to direct light as required for the type of analysis to be carried out. In a preferred embodiment, three types are used, namely, reflective mirrors, dichroic mirrors, and beam splitters, either thin-film beam splitters or partly silvered mirrors Alternatively, they are not used when it is unnecessary to direct the light, e.g. in measuring luminescence and absorbance. A beam dump (an absorber) is provided for use in conjunction with a dichroic mirror or a beam splitter in order to remove excitation light which is not reflected into a sample well and would adversely affect the light detector.
Apertures of various sizes are provided for the excitation light and the emitted light. The selected aperture can be moved into position easily since they are mounted on a moveable plate containing the number of apertures needed for the types of analysis being carried out.
When analyzing a sample by polarization fluorescence, polarizing filters are provided, which filter both the excitation and emitted light either parallel or perpendicular to the polarization direction of the light. In one embodiment, a liquid crystal polarization rotator-linear polarizer combination is used to make it possible to alternate the polarization of the light electronically.
The light emitted by a sample, enters the light detector via a light pipe, i.e. a solid glass pipe, rather than via a lens or multiple group of lenses since imaging of the sample well, or a portion of the sample well is not required.
Light emitted from a sample well may be directed to the light detector from either the top or bottom of the well.
The optical elements are arranged to provide that the excitation light (except for absorbance measurements) has a cross-section equal to the cross-section of the sample well, rather than being limited to a small region within the sample well. Absorbance measurements are made by providing a narrow beam of excitation light at the center line of the sample well. The light emitted from the sample well is not significantly refracted (spread) by the lenses adjacent to the sample well since the emitted light is at the center line of the lenses and collimated.
All measurements of emitted light are made with a single detector in a read head which includes all of the optical elements.
For analysis by absorbance, a diffuser is provided to remove the random polarization caused by excitation light passing through the transparent bottom of a sample well, thus providing more consistent measurements of the emitted light.
The read head remains in one position, while the microplates are positioned horizontally and vertically to accommodate the type of microplate being used.
In one aspect, the invention includes the universal microplate analyzer as configured for analyses by fluorescence, including time-resolved and polarization fluorescence, absorbance, and luminescence.
In another aspect, the analyzer of the invention also includes facilities for carrying out a Luminescent Oxygen Channeling Immunoassay (LOCI), such facilities being designated xe2x80x9cAlpha Screenxe2x80x9d by the Packard Instrument Company and described in U.S. patent application Ser. No. 09/512,707 and incorporated herein by reference.